Functional genomics studies of Salvia miltiorrhiza I. Establish cDNA microarray of S. miltiorrhiza / 中国中药杂志

ABSTRACT

<p><b>OBJECTIVE</b>Establishing cDNA microarray, in order to study functional genomics of Salvia miltiorrhiza.</p><p><b>METHOD</b>Total RNA samples were prepared from S. miltiorrhiza roots using a modified CTAB method. mRNA was isolated by Quichprep Micro mRNA Purification Kit from Pharmacia. Then cDNA was synthesized and cloned into the EcoRI-XhoI sites of the ZAP Express vector using a cDNA synthesis kit, and the ligation mixture was packaged using a ZAP-cDNA Gigapack Gold III cloning kit (Stratagene). The single phage was isolated for PCR amplification, Aliquots of the PCR reactions were analyzed in a 1.5% agarose gel to verify the quality of PCR. The remaining cDNA was purified by Multiscreen filter plates (Millipore) and aliquots were analyzed by agarose gel again to verify the quality of purification. Clones passed verification was resuspended in 15 microL 50% DMSO for arraying. An actin gene from S. miltiorrhiza was used for positive control. PloyA and 50% DMSO was used for negative controls.</p><p><b>RESULT</b>Bacterial colonies containing cNDAs of S. miltiorrhiza were inserted with average insert size of 0. 5 kb to 2. 5 kb. Total 4 354 genes were singled out from the first 8 736 PCR product and used for cDNA microarray manufacture. Single color fluorescence hybridization showed that all positive controls had signals while negative controls had no signals.</p><p><b>CONCLUSION</b>It was the first cDNA microarray about traditional Chinese herbs especially for geoherbs. It could be a powerful tool for studying functional genomics of S. miltiorrhiza.</p>